In vivo calcium imaging reveals directional sensitivity of C-low threshold mechanoreceptors.

C-low threshold mechanoreceptors (C-LTMRs) in animals (termed C-tactile (CT) fibres in humans) are a subgroup of C-fibre primary afferents, which innervate hairy skin and respond to low-threshold punctate indentations and brush stimuli. These afferents respond to gentle touch stimuli and are implicated in mediating pleasant/affective touch. These afferents have traditionally been studied using low-throughput, technically challenging approaches, including microneurography in humans and teased fibre electrophysiology in other mammals. Here we suggest a new approach to studying genetically labelled C-LTMRs using in vivo calcium imaging. We used an automated rotating brush stimulus and von Frey filaments, applied to the hairy skin of anaesthetized mice to mirror light and affective touch. Simultaneously we visualized changes in C-LTMR activity and confirmed that these neurons are sensitive to low-threshold punctate mechanical stimuli and brush stimuli with a strong preference for slow brushing speeds. We also reveal that C-LMTRs are directionally sensitive, showing more activity when brushed against the natural orientation of the hair. We present in vivo calcium imaging of genetically labelled C-LTMRs as a useful approach that can reveal new aspects of C-LTMR physiology. KEY POINTS: C-low threshold mechanoreceptors are sensitive to the directionality of a brush stimulus, being preferentially activated by brushing against the grain of the hair, compared with brushing with the grain of the hair. This is surprising as brushing against the grain of the hair is considered less pleasant. In vivo calcium imaging is a useful approach to the study of C-low threshold mechanoreceptors. While viral transfection, using systemic AAV9, is effective in labelling most sensory neuron populations in the dorsal root ganglion, it fails to label C-low threshold mechanoreceptors.